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| Status |
Public on Aug 04, 2010 |
| Title |
Subtractive cloning of differentially expressed genes in chick heart/hemangioblast precursor cells |
| Organism |
Gallus gallus |
| Experiment type |
Expression profiling by array
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| Summary |
Genetic evidence has implicated several genes as being critical for heart development. However, the inducers of these genes as well as their other targets and the pathways they constitute, remain largely unknown. In the avian embryo, Cerberus (cCer) transcripts are detected in the anterior mesendoderm including the heart precursor cells and in the left lateral plate mesoderm. We have identified a promoter element of chick cCer able to drive EGFP expression into the heart and the hemangioblast precursor cells, allowing us to identify a population of cells that consistently exit from the anterior primitive streak region from as early as stage HH3+ and that later will populate the heart. In order to identify and study novel genes expressed and involved in the correct development and differentiation of the vertebrate H/HPC (Heart/ Hemangioblast Precursor Cells) lineages, a differential screening using Affymetrix GeneChip system technologies was performed. Remarkably, this screening led to the identification of more than 800 transcripts potentially expressed in these precursor lineages. We have identified unknown genes that are differentially expressed in the H/HPC precursor lineages. By developing a procedure to isolate the heart precursor cells using the cer2.5-EGFP construct, we were able to specifically isolate a population of H/HPC expressing already known cardiac markers, and a long list of still uncharacterized genes. From those, we defined by WISH that at least some of these in silico identified transcripts are in fact expressed in these cell populations and more importantly, functionally required for heart formation.More importantly, our study unveiled several uncharacterized genes that can now be used for further studies.
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| Overall design |
Portions of chick embryos were selected and excised at early stages of development for RNA extraction and hybridization on Affymetrix microarrays. We sought to obtain homogeneous populations of embryos at early gastrula stage in order to increase the temporal resolution of expression profiles. To that end, we hand-selected and excised portions of the embryos according to morphological criteria and EGFP expression.
Four GeneChip chicken arrays used for another study were added to the two arrays in this study to ensure a robust model for expression value computation (Note: The 4 samples chick_T2-1, chick_T2-2, chick_T2-3, chick_T2-4, were performed by another group and for a different array, and nothing has to do with our data. They were only run at the same time and used for normalization. Raw data unavailable.). The arrays were normalized to a baseline array with median CEL intensity by applying an Invariant Set Normalization Method. Normalized CEL intensities of the six arrays were used to obtain model-based gene expression indices (MBEI) based on a PM (Perfect Match)-only model.
Processed data for all 6 samples can be found in supplementary file linked below.
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| Contributor(s) |
Bento M, Belo JA |
| Citation missing |
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| Submission date |
Aug 03, 2010 |
| Last update date |
Mar 22, 2012 |
| Contact name |
Margaret Carvalho Bento |
| E-mail(s) |
[email protected]
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| Organization name |
University of Algarve
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| Department |
Biomedical and Medicine
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| Lab |
3.29 Embryology and Genetics
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| Street address |
lab 3.29, Ed 8
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| City |
Gambelas |
| State/province |
Faro |
| ZIP/Postal code |
8005-139 |
| Country |
Portugal |
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| Platforms (1) |
| GPL3213 |
[Chicken] Affymetrix Chicken Genome Array |
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| Samples (2) |
| GSM573672 |
PPPs of embryo at stage HH4-4+, biological rep1 |
| GSM573673 |
HFRs of embryo at stage HH4-4+, biological rep1 |
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| Relations |
| BioProject |
PRJNA131261 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE23389_RAW.tar |
6.3 Mb |
(http)(custom) |
TAR (of CEL) |
| GSE23389_complete_processed_data.txt.gz |
1.0 Mb |
(ftp)(http) |
TXT |
| Processed data included within Sample table |
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